Nova Patents
US8962294B2

Perhydrolase variant providing improved specific activity

Claim Score by NHIP

Read claim 6, the broadest

Abstract

An acetyl xylan esterase variant having perhydrolytic activity is provided for producing peroxycarboxylic acids from carboxylic acid esters and a source of peroxygen. More specifically, a Thermotoga maritima acetyl xylan esterase gene was modified using error-prone PCR and site-directed mutagenesis to create an enzyme catalyst characterized by an increase in specific activity. The variant acetyl xylan esterase may be used to produce peroxycarboxylic acids suitable for use in a variety of applications such as cleaning, disinfecting, sanitizing, bleaching, wood pulp processing, and paper pulp processing applications.

US8962294B2, drawing sheet 1
Sheet 1 of 30

Term

6.9 yearsleft in the term

Expires 14 August 2033, including 296 days of term adjustment.

  1. Priority
  2. Filed
  3. Granted
  4. Today
  5. Expires

24 claims: 2 independent, 22 dependent

  1. 1
    An isolated nucleic acid molecule selected from the group consisting of:(a) a polynucleotide encoding a polypeptide having perhydrolytic activity, said polypeptide comprising the amino acid sequence of SEQ ID NO: 14;(b) a polynucleotide comprising the nucleic acid sequence of SEQ ID NO: 42;and (c) a polynucleotide fully complementary to the polynucleotide of (a) or (b).
  2. 6
    Broadest claimClaim Score 94, very broad(NHIP)An isolated polypeptide having perhydrolytic activity comprising the amino acid sequence of SEQ ID NO:14.
  3. 7
    A process for producing a peroxycarboxylic acid comprising:(a) providing a set of reaction components comprising: (1) at least one substrate selected from the group consisting of: (i) one or more esters having the structure [X] m R 5 wherein X=an ester group of the formula R 6 —C(O)O;R 6 =an unsubstituted C1 to C7 linear, branched or cyclic hydrocarbyl, a substituted C1 to C7 linear, branched or cyclic hydrocarbyl substituted with hydroxyl groups or C1 to C4 alkoxy groups, or comprises C2 to C7 and one or more ether linkages;R 5 =an unsubstituted a C1 to C6 linear, branched, or cyclic hydrocarbyl, or a five- or six-membered heterocyclic aromatic moiety, a substituted C1 to C6 linear, branched, or cyclic hydrocarbyl or a five- or six-membered heterocyclic aromatic moiety substituted with hydroxyl groups;wherein each carbon atom in R 5 individually comprises no more than one hydroxyl group or no more than one ester group or carboxylic acid group or comprising C2 to C6 and one or more ether linkage;m is an integer ranging from 1 to the number of carbon atoms in R 5 ;and wherein said esters have solubility in water of at least 5 ppm at 25° C.;(ii) one or more glycerides having the structure wherein R 1 =an unsubstituted C1 to C21 straight chain or branched chain alkyl or a substituted C1 to C21 straight chain or branched chain alkyl substituted with an hydroxyl or a C1 to C4 alkoxy group;and R 3 and R 4 are individually H or R 1 C(O);(iii) one or more esters of the formula: wherein R 1 =an unsubstituted C1 to C7 straight chain or branched chain alkyl or a substituted C1 to C7 straight chain or branched chain alkyl substituted with an hydroxyl or a C1 to C4 alkoxy group and R 2 is a C1 to C10 straight chain or branched chain alkyl, alkenyl, alkynyl, aryl, (CH 2 CH 2 O) n , or (CH 2 CH(CH 3 )—O) n H and n is 1 to 10;(iv) one or more acylated monosaccharides, acylated disaccharides, or acylated polysaccharides;and (v) any combination of (i) through (iv);(2) a source of hydrogen peroxide;and (3) an enzyme catalyst comprising the polypeptide of claim 6 ;and (b) combining the set of reaction components under suitable reaction conditions whereby peroxycarboxylic acid is produced.
  4. 18
    A composition comprising:(a) a set of reaction components comprising: (1) at least one substrate selected from the group consisting of: (i) one or more esters having the structure [X] m R 5 wherein X=an ester group of the formula R 6 —C(O)O;(i) one or more esters having the structure [X] m R 5 wherein X=an ester group of the formula R 6 —C(O)O;R 6 =an unsubstituted C1 to C7 linear, branched or cyclic hydrocarbyl, a substituted C1 to C7 linear, branched or cyclic hydrocarbyl substituted with hydroxyl groups or C1 to C4 alkoxy groups, or comprises C2 to C7 and one or more ether linkages;R 5 =an unsubstituted C1 to C6 linear, branched, or cyclic hydrocarbyl, or a five- or six-membered heterocyclic aromatic moiety, a substituted C1 to C6 linear, branched, or cyclic hydrocarbyl or a five- or six-membered heterocyclic aromatic moiety substituted with hydroxyl groups;wherein each carbon atom in R 5 individually comprises no more than one hydroxyl group or no more than one ester group or carboxylic acid group or comprising C2 to C6 and one or more ether linkage;m is an integer ranging from 1 to the number of carbon atoms in R 5 ;and wherein said esters have solubility in water of at least 5 ppm at 25° C.;(ii) one or more glycerides having the structure wherein R 1 =an unsubstituted C1 to C21 straight chain or branched chain alkyl or a substituted C1 to C21 straight chain or branched chain alkyl substituted with an hydroxyl or a C1 to C4 alkoxy group;and R 3 and R 4 are individually H or R 1 C(O);(iii) one or more esters of the formula: wherein R 1 =an unsubstituted C1 to C7 straight chain or branched chain alkyl or a substituted C1 to C7 straight chain or branched chain alkyl substituted with an hydroxyl or a C1 to C4 alkoxy group and R 2 is a C1 to C10 straight chain or branched chain alkyl, alkenyl, alkynyl, aryl, (CH 2 CH 2 O) n , or (CH 2 CH(CH 3 )—O) n H and n is 1 to 10;(iv) one or more acylated monosaccharides, acylated disaccharides, or acylated polysaccharides;and (v) any combination of (i) through (iv);(2) a source of hydrogen peroxide;and (3) an enzyme catalyst comprising the polypeptide of claim 6 ;and (b) at least one peroxycarboxylic acid formed upon combining the set of reaction components of (a).
Independent claims4